Alkylated cortical steroids



United States Patent Ofiice 3,053,832 ALKYLATED CORTICAL STEROIDS DavidH. Gould, Leonia, N.J., assignor to Schering Corporation, Bloomfield,N.J., a corporation of New Jersey No Drawing. Filed Apr. 29, 1957, Ser.No. 655,476 12 Claims. (Cl. 260-2395) This invention relates to a newgroup of corticoid steroids and particularly to the 6oc-methyl analogsof prednisone (1,4 pregnadiene-17a,21-diol-3,l1,20-trione), prednisolone(1,4 pregnadiene 11 ,8,17a,21-triol-3,20-dione), their 9oc-fll1010derivatives and the 2l-esters thereof.

It is an object of this invention to provide steroid compounds havingthe aforementioned description which exhibit modified adrenocorticalactivity, ie more selective effects with a reduction or minimum ofadverse side reaction. It is a further object of this invention toprovide processes for the manufacture of these valuable steroidcompounds.

The discovery of the remarkable properties of prednisone andprednisolone has been recognized as a major step forward in thetreatment of arthritis and related diseases. It is now known thatprednisone and prednisolone provide definite advantages over cortisoneand bydrocortisone, both in their degrees of potency and exhibition ofside effects. The principal advantage which has been realized with theadvent of prednisone and prednisolone aside from potency, has been areduction in mineralocorticoid activity, i.e. a decrease in saltretention. Despite the fact that the salt retention elfects ofprednisone and prednisolone are remarkably reduced in comparison withcortisone and hydrocortisone, there has still been room for improvement.There has been a continuing Widespread interest in finding othersteroids and derivatives which would perhaps be more potent thanprednisone and prednisolone, but at least exhibit even less saltretaining effects. I have found that the 6a-methyl prednadienes,described above, advantageously possess such properties. Thenewcompounds of my invention may be represented by the following generalformula:

wherein X is a member of the group consisting of O and (H,fiOH), Y is amember of the group consisting of H and F, and R is a member of thegroup consisting of H and acyl.

The new compounds of the formula may be used as the free 21-alcohol, forexample, when oral administration is I lower doses for a given set ofindications. Representative of the esters, which are advantageous, arethe lower alkanoyl esters such as acetate, propionate, trimethylacetate, cyclopentylacetate, phenoxyacetate, furoate and the like. Inaddition to esters with mono-basic organic acids such as describedheretofore, it is sometimes advantageous to prepare esters of diorpoly-basic acids so as to provide a free acid group which upon reactionwith base would render the steroid ester soluble in Water or aqueoussolutions. For example, a common lower alkanoyl di-basic acid used inthis respect is succinic acid. The hemisuccinate of the steroid isprepared in a conventional manner and the ester is solubilized in theform of its sodium salt. Poly-basic inorganic acids also provide forwater solubility, as particularly evidenced by a 2l-phosphate ester.After having formed a mono-ester between the sterol and the phosphoricacid in a manner well-known in the art, the resultant 21phosphate issolubilized generally in the form of its sodio derivative. The sodiumsalt is soluble in a large range of solvents including water, andmaintains the same high cortical activity as does the free sterol.

The compounds of my invention are prepared by first reacting a3-keto-4-pregnene precursor (I) with a secondary amine so as to form anene-amine. More specifically, the starting material may be cortisone,hydrocortisone, their 9a-fluoro analogs or their 21-esters, which uponreaction with a cyclic secondary amine forms a 3-amino-3,5-pregnadieneintermediate (II). In order to properly carry out the synthesis of thecompounds of this new invention, especially the step requiringalkylation at C-6, it is necessary that careful selection be made in thechoice of secondary amine. I prefer to use a cyclic amine which issterically hindered, such as Z-methyl piperidine or preferably2,6-dimethyl piperidine. The presence of the methyl groups in closeproximity to the nitrogen atom effectively cages vulnerable points ofattack in the ene-amine (II) molecule. Specifically, alkylation isdirected towards C-6 while alkylation of the nitrogen atom or of C-4 isdiscouraged by the presence of the methyl groups. I have further foundthat it is preferable to remove the water formed from the reactionbetween the 3-keto-4-pregnene and the piperidine so as to displace theequilibrium in the desired direction.

The crude ene-amine (II) may be alkylated merely by refluxing with amethylating agent such as methyl iodide giving rise to a 6-metl1ylene-amine (III). Hydrolysis of III with mild aqueous acid or baseregenerates the 3-keto- M-pregnene thereby forming the Got-methylderivative of cortisone, hydrocortisone or their 9u-fluoro analogs ortheir 2l-esters (IV).

The 6a-methyl-4-pregnene (IV) is preferably first hydrolyzed and thendehydrogenated so as to form a 1,4 prcgnadiene according to methods nowwell-known in the art. I prefer to employ microbiological technics suchas subjecting the 6amethyl-4-pregnene to the dehydrogenating action ofBacillus sphaericus (A.T.C.C. 7055) whereby the diene (V) is easilyformed. A compound of Formula V may be purified at this stage andcompounded pharmaceutically for therapeutic application. If an ester isdesired, said ester may be prepared ina conventional manner forpreparing 21-esters such as by re-.

acting the Zlalcohol with an acylating agent such as an anhydride oracid chloride in a pyridine solution. Esters such as hemi-succinates andphosphates are conveniently Patented Sept. 11., I962 prepared accordingto the techuics fully described in the chemical and patent literature.

The following equation schematically sets forth the foregoingtransformations:

CHzOR methyletion O H] CHaOR CHzOR X i 1) hydrolysis 2) microbiologicaldehydrogenation V CH;

The following examples illustrate methods for preparing the newcompounds of my invention. It is to be understood, however, that myinvention is limited only as defined in the appended claims.

4 EXAMPLE 1 9a-Fluoro-6a-Methyl-1,4-Pregnadiene-1 113,1 7u,21Trial-3,20-Di0ne A. 3 -(2',6-DIMETHYLPIPERIDINO)- 9a FLUORO 3,6-PREGNADIENE-l1B,17a,21-TRIOL-20'ONE 21-ACETA'IE Nitrogen gas is bubbledthrough a solution of 2 grams 9u-fluorohydrocortisone acetate, inethylene dichloride. There is then added 0.55 g. of2,6-dimethylpiperidine (1.05 equiv.) and 0.02 g. of p-toluene sulfonicacid. The solution is refluxed for two hours in an apparatus whichpermits the refluxing liquid to return through calcium carbide (toremove Water) before returning to the reaction vessel. The reactionmixture is then treated with 0.01 g. of sodium bicarbonate andconcentrated to dryness in vacuo. The ene-amine so obtained is useddirectly in the next step.

B. 9a-FLUORO-6a-l\IETHYL-4.PREGNENE-11B,17a,21-

TRIOL-3,20DIONE 21-ACETA1E One gram of the intermediate ene-amineprepared above, is dissolved in 20 ml. of dry acetone and 20 ml. ofmethyl iodide. The solution is refluxed for six hours. After evaporationof the solvent, the residue is dissolved in 20 ml. of acetic acid towhich is added 2 g. of sodium acetate and the mixture is refluxed for 2hours. After cooling, the mixture is diluted with water and extractedtwice with methylene chloride. The extracts are combined, washed oncewith sodium carbonate-solution and then with Water until the waterwashes are neutral. The methylene chloride solution is then dried oversodium sulfate, filtered, and concentrated to a small volume. Hexane isadded, then this mixture is poured onto a column of activated magnesiumsilicate. The column is washed first with hexane, then elution iscontinued with dry ether in hexane. The fraction eluted with 50-75%ether in hexane is crystallized from acetone-hexane to give9u-fluoro-6a-methyl-4-pregnene-11fi,l7a,21-triol-3, 20-dione 21-acetate.

C. 9a-FLUORO-Ga-METHYL4-PREGNENE-11B,17a,21-

TRIOL-3,20-DIONE A sample of 0.37 g. of the product of Procedure B isdissolved in 13 ml. of 1:2 chloroform-methanol mixture, cooled to 15 C.,and stirred under argon for ten minutes. Then, with continued stirring,there is quickly added 0.86 ml. of an aqueous solution containing 35 mg.of sodium hydroxide. Stirring is continued for two minutes, then 0.2 ml.of acetic acid is added, and the reaction mixture is concentrated invacuo to dryness. The residue is suspended in warm water, filtered, anddried in a vacuum oven at 75 C. The dried solid, which is 90:- fiuoro 6amethyl 4 pregnene 1lB,17a,21 triol 3, 20-dione, is recrystallized fromacetone.

D. Qa-FLUORO-Ga-METHYLI,4-PREGNADIENE-11fl,17a,21-

TRIOL-3,20-DIONE A mixture of 10 g. of yeast extract (Difco) in l 1. oftap water, the pH of which is adjusted to 6.8-7.0, is distributed amongten 300-ml. Erlenmeyer flasks and to each flask is added an inoculum ofB. sphaericus. The resulting suspensions are incubated at 30C. on ashaking machine and growth is followed turbidimetrically. The startingmaterial (0.1 g. of the product of Procedure C) is dissolved in 10 m1.of methanol. This solution is distributed equally among the ten flaskswhen the peak of the log phase of growth of the organism is reached.Progress of the reaction is then followed by withdrawing flasks from theshaking machine at periodic intervals, extracting the mixture withchloroform and chromatographing the extracted steroids according to themethod outlined by G. M. Schull on page 9A of Abstracts of Papers, 126thMeeting of the Am. Chem. Soc., September 12-17, 1954, New York, NewYork. When the desired transformation is complete (3-48 hours), as shownby the disappearance of the starting material or the absence of furtherchange in the composition of the reaction mixture, the contents of theremaining flasks are combined and extracted with chloroform. The crudechloroform extract from the transformation is concentrated to a residuewhich is crystallized from methylene chloride-hexane to give9a-fiuoro-6a-methy1-1,4-pregnadiene-l1,6,17a,21-triol-3,20-dione.

EXAMPLE 2 9a-Fluor0-6a-Methyl-1 ,4-Pregnaaz'ene-115J 70:,21-

Trial-3,20-Dine 21-Acetate EXAMPLE 3 To a sample of 50 mg. of theproduct of Example 2 suspended in ml. of glacial acetic acid, is added14 mg. of chromic anhydride dissolved in 1 ml. of water. The mixture isstirred at room temperature for four hours during which time the solidgoes into solution. The solution is then poured into 15 ml. of watercontaining 50 mg. of sodium sulfite, and allowed to stand one hour. Theproduct, 9a-fluoro-6a-methyl-1,4pregnadiene-17a,21- diol-3,11,20-trione21 acetate, is filtered oft, dried and crystallized from acetone-hexane.

EXAMPLE 4 6 u-M ethylprednisone A. 3-(2'-METHYLPIPERIDINO)-3,5PREGNADIENE- 17a,21-DIOL-l1,20-DIONE 21-DIETHYLACETATE' Usingprocedure A of Example 1, 2 grams of cortisone 2l-diethylacetate (M.P.167-l69 C.) are treated with 0.50 g. of Z-methylpiperidine. The finalresidue, which is the ene-amine of this example, is used directly in thenext step.

B. Ga-METHYLCORTISONE DIETHYLACETATE Using procedure B of Example 1, 2grams of the intermediate ene-arnine prepared above are treated with 40ml. of methyl iodide in 40 ml. of dry dimethylformamide. Theintermediate is hydrolyzed with aqueous acetic acid and chromatographed.The fraction eluted with 30-50% ether in hexane is crystallized frommethanol to give 6a-methylcortisone diethylacetate.

C. Ga-METHYLCORTISONE One gram of the 6a-methylcortisone diethylacetateprepared above is treated with 90 mg. of sodium hydroxide for fiveminutes according to procedure C of Example 1. The crude solid iscrystallized from methanol to yield 6a-methylcortisone.

D. Ga-METHYLPREDNISONE Procedure D of Example 1 is repeated using assubstrate in each flask 50 mg. of the 6a-methylcortisone prepared above.The combined extracts are concentrated and crystallized fromacetone-hexene to yield 6u-methylprednisone.

EXAMPLE 5 6 oc-M ethylprednisolone A. 3- (2',6-DIMETHYLPIPERIDINO-3,5-PREGNADIENE- 1 1B,17a,2l-TRIOL-ZO-ONE ZI-ACETATE Procedure A ofExample 1 is repeated using 2 g. of hydrocortisone acetate and 0.59 g.of 2,6-dimethylpiperidine. The semi-crystalline residue, the ene-amineintermediate, is used directly in the next step.

B. Ba-METHYLHYDROCORTISONE 21-ACETATE The intermediate ene-amine,prepared above, is treated according to procedure B of Example 1. Thecrude product is chromatographed, and the fraction eluted with 25-0%hexane in ether is crystallized from acetonehexane to yield6a-methylhydrocortisone 21-acetate.

C. Ba-METHYLHYDROCORTISONE The 21-acetate ester prepared above issaponified as in procedure C of Example 1. The precipitate is crystallized from acetone-hexane to yield 6a-methylhydrocortisone.

D. (ta-METHYLPREDNISOLONE The 6a-methylhydrocortisone prepared above istreated as in procedure D of Example 1 to give a residue which iscrystallized from methanol to yield the compound of this example,6a-methylprednisolone.

EXAMPLE 6 6 a-M ethylprednisolone 21 -A cetate The product of Example 5is acetylated as in Example 2. The precipitate is crystallized frommethylene chloride-hexane to yield 6a-methylprednisolone 21-acetate.

I EXAMPLE 7 6a-Methylprednis0ne 21 -Acetate The product of Example 6 isoxidized as described in Example 3. The crude product is recrystallizedfrom methanol to yield 6a-methylprednisone 21-acetate.

I claim:

1. A process which comprises reacting a 3-keto-4- pregnene selected fromthe group consisting of 4-pregnene 17a,21 diol-3,11,20-trione,4-pregnene-11;8,17a,21- triol-3,20-dione,9ot-fluoro-4-pregnene-17a,21-diol-3,l1,20- trione, c fluoro4-pregnene-115,17a,21-triol-3,20-dione and the 21-esters thereof with acyclic secondary amine having a methyl group on a carbon atom adjacentto the nitrogen 'atom to cause the formation of a 3-amino-3,5-pregnadiene; alkylating said pregnadiene at the 6-carbon position;hydrolyzing said pregnadiene to cause the regeneration of the3-keto-4-pregnene; and subjecting this latter product to microbiologicaldehydrogenation with Bacillus sphaericus to cause the production of thecorresponding 3-keto-l,4-pregnadiene alkylated at the 6-carbon position.

2. A process which comprises reacting a 3-keto-4- pregnene selected fromthe group consisting of 4-pregnene :,21 diol-3,11,20-trione,4-pregnene-1l/3,l7a,21- triol-3,20-dione,9a-fluoro-4-pregnene-17a,21-diol-3,l1,20- trione, 90c fluoro4-pregnene-1 1,8,17a,21-triol-3,20-dione and the 2l-esters thereof witha cyclic secondary amine having a methyl group on a carbon atom adjacentto the nitrogen atom to cause the formation of the corresponding3-amino-3,5-pregnadiene of said 3-keto-4-pregnene and said cyclic amine;alkylating said 3-amino-3,5-pregnadiene to produce the corresponding3-amino-6-methyl- 3,5-pregnadiene; hydrolzing said pregnadiene to causethe preparation of the corresponding 6-methyl-3-keto-4-pregnene selectedfrom the group consisting of 4-pregnene- 1704,21 diol 3,11,20-trione,4-pregnene1l,8,l7a,2l-triol- 3,20 dione, 9a fluoro4-pregnene-17a,21-diol-3,11,20- trione, 9a fluoro4-pregnene-115,17a,21-triol 3,20-dione and the 2l-esters thereof andsubjecting said 6-methyl-4- pregnene to the microbiologicaldehydrogenative activity of Bacillus spha ericus to cause the formationof the corresponding 1,4-diene of said 3-keto-4-pregnene.

3. A process which comprises reacting a 3-keto-4-pregnene selected fromthe group consisting of 4-pregnene- 17a,21-dio1-3,11,20-trione,4-pregnene-11/3,17a,21-triol-3, 20-dione,9a-fiuoro-4-pregnene-17a,21-diol-3,11,20-trione,9a-fluoro-4-pregnene-11,8,17a,21-triol-3,20-dione and the 21-estersthereof with a cyclic secondary amine having a methyl group on a carbonatom adjacent to the nitrogen atom to cause the formation of thecorresponding 3- amino-3,5-pregnadiene of said 3-keto-4-pregnene andsaid cyclic amine; alkylating said 3-amino-3,5-pregnadiene to producethe corresponding 3-amino-6-methyl-3,5-pregnadiene; hydrolyzing saidpregnadiene to cause the preparation of the corresponding6-rnethy1-3-keto-4-pregnene selected from the group consisting of4-pregnene-17u,2ldiol 3,11,20 trione, 4 pregnene 11B,17oc,21 triol- 3,20dione, 90c fluoro 4 pregnene 170:,21 diol- 3,11,20 triol, 9a fluoro 4pregnene 11fi,17a,21- triol-3,20-dione and the 21-esters thereof.

4. A process which comprises reacting a 3-keto-4-pregnene selected fromthe group consisting of 4-pregnene- 170:,21 diol 3,11,20 trione, 4pregnene 115,111,21- triol 3,20 dione, 9oz fluoro 4 pregnene 170:,21-diol 3,11,20 trione, 90c fluoro 4 pregnene 11/3, 1711,21 triol 3,20dione and the 21-esters thereof with a cyclic secondary amine having amethyl group on a carbon atom adjacent to the nitrogen atom to cause theformation of the corresponding 3-amino-3,5-pregnadiene derivativethereof; and alkylating said 3-amino-3,5-pregnadiene derivative toproduce the corresponding S-amino- 6-methyl-3,5-pregnadiene.

5. A process which comprises reacting a 3,5-pregnadiene selected fromthe group consisting of 3-X-3,5- pregnadiene 17a,2l diol 11,20 dione,3-X-3,5-pregnadiene 1lfl,17a,2l triol 20 one, 3-X-9a fluoro 3,pregnadiene 17a,21 diol 11,20 dione,3-X-9ocfluoro-3,5-pregnadiene-11,3,17a,21-triol-20-one and the 21-estersthereof wherein X is a cyclic secondary amine substituent having amethyl group on a carbon atom adjacent to the nitrogen atom with amethylating agent to cause the formation of the corresponding6-methyl-3- amino-3,5-pregnadiene.

6. A process which comprises reacting 3-(2,6'-dimethylpiperidino) 9afiuoro 3,5 pregnadiene 11,9,17a, 21-triol-20-one with methyl iodide toyield the corresponding 6-rnethyl ene-amine thereof.

7. A process which comprises reacting 3-(2-methyl piperidino) 3,5pregnadiene l7a,21 diol 11,20- dione 21-dietl1ylacetate withmethyliodide to yield the corresponding 6-methyl ene-amine thereof.

8. A process which comprises reacting 3-(2',6'-dimethylpiperidino) 3,5pregnadiene 11;3,17a,21 triol 20- one 21-acetate with methyliodide toyield the corresponding 6-methyl ene-amine thereof.

9. 3 (.2',6 dimethylpiperidino) 6 methyl 9afluoro 3,5 pregnadiene1lfl,l7a,21 triol 20 one 2l-acetate.

10. 3 (2 methylpiperidino) 6 methyl 3,5 pregnadiene 1701,21 diol 11,20dione 21 diethylacetate.

11. 3 (2',6' dimethylpiperidino) 6 methyl 3,5-pregnadiene-l1,3,l7a,21-triol-20-one 21-acetate.

12. A compound of the formula:

CHzOR I =0 133 a f i References Cited in the file of this patent UNITEDSTATES PATENTS 2,781,342 Herr Feb. 12, 1957 2,781,343 Magerlein Feb. 12,1957 2,852,511 Fried Sept. 16, 1958 2,897,218 Sebek July 28, 1959 OTHERREFERENCES Stoudt, Chem. Abstracts, page 3558, March 10, 1956.

Spero et al.: J.A.C.S. 78, December 5, 1956, pages 6213, 6214.

12. A COMPOUND OF THE FORMULA: